Isothermal titration microcalorimeter apparatus and method of use

ABSTRACT

An automatic pipette assembly for an isothermal titration micro calorimetry system, comprising a pipette housing, a syringe with a titration needle arranged to be inserted into a sample cell for supplying titrant, and a linear activator for driving a plunger in the syringe, the titration needle is rotatable with respect to the housing and is provided with a stirring paddle arranged to stir fluid in the sample cell, wherein the automatic pipette assembly comprises a stirring motor for driving the rotation of the titration needle. There is also provided an isothermal titration micro calorimetry system.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. provisional patent applicationNo. 60/984,432 filed Nov. 1, 2007; the disclosure of which isincorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates generally to microcalorimeters and morespecifically to features that improve the performance ofmicrocalorimeters, especially isothermal titration calorimeters.

2. Background

Microcalorimeters are broadly utilized in fields of biochemistry,pharmacology, cell biology, and others. Calorimetry provides a directmethod for measuring changes in thermodynamic properties of biologicalmacromolecules. Microcalorimeters are typically two cell instruments inwhich properties of a dilute solution of test substance in an aqueousbuffer in a sample cell are continuously compared to an equal quantityof aqueous buffer in a reference cell. Measured differences between theproperties of the two cells, such as temperature or heat flow, areattributed to the presence of the test substance in the sample cell.

One type of microcalorimeter is an isothermal titration calorimeter. Theisothermal titration calorimeter (ITC) is a differential device, butoperates at a fixed temperature and pressure while the liquid in thesample cell is continuously stirred. The most popular application fortitration calorimetry is in the characterization of the thermodynamicsof molecular interactions. In this application, a dilute solution of atest substance (e.g., a protein) is placed in the sample cell and, atvarious times, small volumes of a second dilute solution containing aligand, which binds to the test substance, are injected into the samplecell. The instrument measures the heat which is evolved or absorbed as aresult of the binding of the newly-introduced ligand to the testsubstance. From results of multiple-injection experiments, properties,such as, the Gibbs energy, the association constant, the enthalpy andentropy changes, and the stoichiometry of binding, may be determined fora particular pairing between the test substance and the ligand.

While currently utilized ITCs provide reliable binding data results,their widespread utilization in the early stages of drug developmenthave been limited by several factors: the relatively high amounts ofprotein required to perform a binding determination (e.g., about 0.1milligram (mg) to about 1.0 mg of a protein), the time required toperform the measurement, and the complexity of using conventional ITCs.Due to the extremely high costs of biological substances used inresearch, there is a need to reduce the amount of biological substanceused for each experiment. A reduction in the amount of the biologicalsubstance used in a calorimeter experiment, will require a moreaccurate, sensitive, and reliable titration calorimeter than what iscurrently available.

Furthermore, gathering binding data utilizing prior art ITCs requireextensive preparation and skill by the practitioner. For example, usingprior art ITCs, the reference and sample cells are first filledrespectively with the reference substance and sample substance via acorresponding cell stem. Next, a syringe of the ITC is filled with atitrant. Then a needle of the ITC is placed in the sample cell via acell stem leading to the sample cell while the syringe fits into aholder on the ITC enabling the syringe to rotate around its axis.Subsequently, the syringe is aligned with the sample cell so that theneedle does not touch either the cell stem or the sample cell. Then, thesyringe is connected to a stirring motor and a linear activator of theITC, wherein the stirring motor and the linear activator must also bealigned with the sample cell.

As would be appreciated by a reading of the above-described prior artprocedure, utilizing prior art ITCs, the quality of binding measurementsperformed with these prior art ITCs depends heavily of the operator'sskills and experience, and involves a considerable amount of preparationtime.

More recently developed prior art ITCs have attempted to simplify thepreparatory procedures described above. For example, in one such priorart ITC, which is partly shown in FIG. 1 as ITC 100, a syringe 102, asyringe holder 104, and a linear actuator 106, which actuates syringe102's plunger, are integrated into a single unit referred to as anautomatic pipette. ITC 100 further comprises a stirring mechanismcomprising a stirring motor 108, which is attached to calorimeter body110. ITC 100 also comprises an inner magnet couple 112 located aroundsyringe 102, and an outer magnet couple 114 located on calorimeter body110 in close proximity to stiffing motor 108. The rotation from stirringmotor 108 to syringe 102 is transferred via magnet couplings 112 and114. Attached to syringe 102 is a needle 116 and a paddle 118. Theneedle 116 is arranged to be inserted into a sample cell 120 via a cellstem 122 for performing ITC experiments. For reference purposes, ITC100, also comprises a reference cell, not shown, in communication withthe ambient atmosphere via a reference cell stem.

The prior art design discussed above and depicted in FIG. 1 has certainlimitations. For example, since the magnet coupling is a soft/flexibletransmission, it is prone to resonant vibration of the stirrer atcertain rotation speeds and accelerations, which negatively affects theinstrument's sensitivity. The resonant vibration can be reduced byeither employing a less sensitive feedback mechanism controlling therotation speed (which leads to less stable rotation speed), or bylowering the rotation speed. However, less stable rotation speed alsoreduces the ITC's sensitivity, while lower rotation speed impedes propermixing of reagents which reduces the ITC's accuracy.

Another limitation of the prior art design is that the stirring motorand the magnet coupling are placed closely to the sensitive measuringunit of the instrument and generates a substantial alternating magneticfield that produces electric noise which negatively affects theoperation of the ITC's sensitive electronic circuitry. Since the ITC'ssensors process signals of approximately 10⁻⁹ volts, and the noisegenerated by the motor and the magnetic coupling is a reciprocal of thedistance between the sensor and the source of the noise, furtherimprovements in the performance characteristics of this ITC designbecome increasingly challenging. As stated earlier, one of theunderlying factors affecting the design of new microcalorimeters is theneed to reduce the amount of biological substance used for eachexperiment. This requires smaller sample cells and shorter cell stemswhich in turn leads to, smaller distances between the cell sensor andthe motor and magnetic coupling (source of electric noise), which limitsthe instrument's sensitivity.

The invention described herein is aimed to improve the aforementionedcharacteristics and use of prior art ITCs such that the sensitivity ofthe ITC is improved, the amount of biological substance necessary fortesting is reduced, the reliability of the results generated by the ITCis improved, and use of the ITC is eased.

SUMMARY OF THE INVENTION

The object of the invention is to provide a new automatic pipetteassembly for an isothermal titration micro calorimetry system and an ITCsystem, which pipette assembly and ITC system overcomes one or moredrawbacks of the prior art. This is achieved by the pipette assembly andthe ITC system as defined in the independent claims.

One advantage with the present pipette assembly and the associated ITCsystem is that it makes it possible to reduce the cell compartmentvolume by about a factor of seven as compared to prior art ITCs, withouta reduction in sensitivity, and with a significantly faster responsetime. Such an ITC system permits the performance of experiments withabout 10 times less protein sample, and with only a total of about 2 toabout 4 titrations per hour.

In addition to reducing the costs associated with running the ITCexperiment, a smaller cell volume also extends the number of ITCapplications. For example, the range of binding affinities that can bemeasured by ITC is dictated by a parameter called “c value,” which isequal to the product of the binding affinity (K_(a)) and the totalconcentration (M_(total)) of macromolecule (c=[M_(total)]K_(a)). Foraccurate affinity determination, the c value must be between 1 and1,000. A decrease in the cell volume by a factor of ten results in asimilar increase in c value if the same amount of protein is used, and,consequently, the ability to measure weak binders. This ability isespecially important in the early stages of drug discovery, in whichbinding affinities are weak, especially in conjunction with a fullyautomated instrument.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a schematic example of a prior art isothermal titrationmicrocalorimeter highlighting the automatic pipette feature of themicrocalorimeter;

FIG. 2 shows, in cross-section, a schematic embodiment of an isothermaltitration calorimeter system of the present invention with a guidingmechanism;

FIG. 3 shows, in cross-section, a schematic embodiment of an automaticpipette assembly for an isothermal titration micro calorimetry system;

FIG. 4 shows a section of the ITC system shown in FIG. 2, wherein thepipette assembly is retracted from the sample cell;

FIGS. 5 a to 5 c show a schematic embodiment of a guiding mechanism foran isothermal titration calorimeter system in perspective;

FIG. 6 shows another schematic embodiment of a guiding mechanism for anisothermal titration calorimeter system in perspective.

DETAILED DESCRIPTION OF THE INVENTION

FIG. 2 schematically shows one embodiment of an ITC system 200 accordingto the present invention. The ITC system 200 comprises a microcalorimeter 210 and an automatic pipette assembly 220. The microcalorimeter 210 comprises a reference cell 240 and a sample cell 250which are designed to be essentially identical in heat capacity andvolume. The cells 240 and 250 are comprised of a suitable chemicallyinert and heat conductive material, such as gold, Platinum, tantalum,hastelloy or the like. The cells 240 and 250 may be of essentially anysuitable shape, but it is desirable that they are of the same shape,that they are possible to arrange in a fully symmetric arrangement, andthat efficient mixing of the titrant with the sample may be achieved. Inthe disclosed embodiment, the cross-section of the cells 240 and 250 isrectangular, and the cross-section in the transverse horizontaldirection may be circular, resulting in coin shaped cells with circularfacing surfaces.

In order to reduce any external thermal influences to a minimum, the,reference cell 240 and the sample cell 250 are both enclosed by a firstthermal shield 260 which in turn is enclosed by a second thermal shield270. The thermal shields 260, 270 may be comprised of any suitablethermally conductive material such as silver, aluminum, cupper or thelike. The shields 260, 270 may further be comprised of one or morethermally interconnected sub shields (not shown, to provide even furtherstable temperature conditions for the calorimetric cells 240, 250.

In order to control the temperature of the shields 260, 270, thermalcontrol means may be arranged to control the temperature thereof. In anITC system said thermal control means are mainly used to set the“isothermal” temperature of the calorimeter, ie. of the thermal shields260, 270, before the titration experiments are initiated. But as will bedisclosed in greater detail below, said thermal control means may alsobe used to improve the adiabatic behavior of the calorimeter. Accordingto one embodiment, the thermal control means are comprised of one ormore heat pump unit, such as a thermoelectric heat pump device based onthe peltier effect or the like. Other types of thermal control meansinclude thermostatically controlled liquid baths, mechanical heat pumps,chemical heating or cooling systems or the like.

In the disclosed embodiment a first heat pump unit 280 is arranged totransfer heat energy between the first 260 and second thermal shields270, a second heat pump unit 290 is arranged to transfer heat energybetween the second thermal shield 270 and a heat sink 300 in thermalcontact with the ambient temperature. A temperature controller 310 isarranged to control the first and second heat pump units 280, 290 sothat the desired temperature conditions are achieved. The temperaturecontroller 310 and associated sensors will be disclosed in more detailbelow.

A reference cell stem 320 and a sample cell stem 330 provides access tothe reference cell 240 and sample cell 250, respectively, for supplyingreference and sample fluids, titration fluid, washing of the cells etc.In the disclosed embodiment, the cell stems 320 and 330 both extendsessentially vertically through both thermal shields and the heat sink toprovide direct communication with cells 240 and 250 and the cell stems320 and 330 each support their respective cell 240 and 250 in the cavityof the first thermal shield 260.

The automatic pipette assembly 210 comprises a pipette housing 340, asyringe 350 with a titration needle 360 arranged to be inserted into thesample cell 250 for supplying titrant, and a linear activator 370 fordriving a plunger 380 in the syringe 350. The titration needle 360 isrotatable with respect to the housing 340 and is provided with astirring paddle 390 arranged to stir sample fluid in the sample cell 250in order to achieve efficient mixing of titrant and sample fluid. Theautomatic pipette assembly 210 further comprises a stirring motor 400for driving the rotation of the titration needle 360.

In the embodiment disclosed in FIG. 2 the stiffing motor 400 is a directdrive motor with a hollow rotor 410 arranged concentric with the syringe350 and the titration needle 360. The syringe 350 is at its upper endsupported for rotation by the stiffing motor 400 and at the lower end bya bearing 420. Both the stirring motor 400 and the bearing 420 areschematically disclosed as comprising ball bearings, but any other typeof bearing, bushings or the like capable of providing smooth and lowfriction rotation of the titration needle 360 may be used.

In the embodiment disclosed in FIG. 2, the syringe 350 is arranged to berotatable with respect to the housing 340 and the titration needle 360is non-rotatably attached to the syringe 350. In an alternativeembodiment (not shown in figs.), the needle 360 is rotatable withrespect to the syringe 350 and the syringe 350 static with respect tothe pipette housing 340. FIG. 3 shows an alternative embodiment, whereinthe syringe 350 is detachably arranged in a rotation frame 430 withinthe pipette housing 340, the rotation frame 430 being rotably supportedby the motor 400 and bearing 420. In the disclosed embodiment, thesyringe 350 is retained in the rotation frame 430 by a cap 440 that isdetachably attached to the rotation frame 430 by threads or the like. Byremoving the cap 440, the syringe 350 with the titration needle 360 canbe replaced.

In an alternative embodiment, not shown in the figures, the stirringmotor 400 drives the titration needle for rotation by a rotationtransmission arrangement, such as a drive belt arrangement, a drivewheel arrangement or the like. In such an arrangement, the drive motormay be placed at an ever greater distance from the calorimetric cells.

The stirring motor is controlled by a stir controller 450 of the ITCsystem. The stir controller be a conventional BLDC. The linear actuatoris controlled by a titration controller 460 of the ITC system. The stirand titration controllers 450, 460 may be arranged in the pipetteassembly 220, and in turn connected to the ITC control system (not shownin detail), or they may be an integrated part of the ITC control system.

In the disclosed embodiment, the linear activator 370 comprises astepper motor 470 arranged to drive a threaded plunger 380 that extendscoaxially through the hole of a hollow rotor 480 and into the syringe350 wherein it is rotatably attached to a pipette tip 490 that sealsagainst the inner wall of the syringe 350 to allow displacing a precisevolume of titration liquid from syringe 350. The pipette assemblyfurther comprises position sensors 500 a, 500 b for detecting twopredetermined positions of the threaded plunger 380. The linearactivator 370 may be of any other type capable of perform controlledlinear motion with sufficient precision. This design allows syringe tobe rotated independently of the main body 340 of the pipette assembly220; at the same time, the linear activator 370 can drive the threadedplunger 380.

In the ITC system the titration 460 controller is arranged to controlthe linear activator of the pipette assembly 220. According to oneembodiment, titration controller 460 uses the position sensors 500 a,500 b for detecting two predetermined positions of the threaded plunger,and the titration controller 460 is arranged to register the twopredetermined positions of the threaded plunger 380 and to determine thepitch of the threads of the plunger 380 from the number of stepsperformed by the stepper motor 470 to move the threaded plunger 380between said positions. The so determined pitch of the threaded plungeris thereafter used by the pipette controller to increase the accuracy ofthe titration pipette, when displacing small volumes of titrant.

In the disclosed embodiments, the pipette assembly housing 340 serves asmounting base for the stators of the stirring motor 400 and the linearactivator 470. The housing 340 may further comprise an attachmentsection for precise positioning of the pipette assembly with respect tothe sample cell stem.

The automatic pipette assembly 220 with an integrated stiffing motor 400arranged at the upper end of, or above the syringe 350 not onlyincreases the distance between the stirring motor 400 (i.e., the sourceof an alternating electro-magnetic field) and the sensitive electroniccircuitry, which is located at and/or nearby the thermal core of thecalorimeter, but it also allows for the reduction in the amount of powerneeded to operate the stirring motor 400. That is, due to the placementof the stiffing motor 400 on the pipette assembly 220, the size of thestirring motor 400 is not determined by the space between the samplecell and the reference cell as it is determined by prior art ITCs thatplace the stiffing motor on the calorimeter housing (FIG. 2). In theprior art, then, the cell stems protrude through the center of rotationof the magnetically coupled stirrer. Therefore, the lower limit of sizefor the stirrer mechanism is determined by the spacing between the cellstems. Accordingly, the size of the stiffing motor of the presentinvention can be significantly reduced, e.g., by a factor of about 5times that of stiffing motors found in prior art ITCs, thereby resultingin a significant reduction in the amount of power needed to operate theITC, e.g., about 2 watts versus the 100 watts used to powerconventionally known ITCs.

The reduction in size of the stirring motor to a size of about fivetimes less than stiffing motors of prior art ITCs, and the placement ofthe stirring motor on the pipette such that the stirring motor is at adistance of about 50 millimeters or more away from the sensitiveelectronic circuitry of the calorimeter, and the removal of the magneticcoupling, closes the magnetic field that exists in conventionally usedITCs. Accordingly, the sensitivity of the inventive ITC is raised by thelower power, the lower heat, the lower electricity, and the lower noiseand vibration caused by the placement of the stirring motor as disclosedherein. Furthermore, the disclosed embodiments also exclude magneticcoupling which is an additional source of alternating electro-magneticfield.

These improvements reduce the electrical noise induced in the sensitiveelectronic circuitry which, in turn, helps improve the calorimeter'ssensitivity, whereby the volume of the sample and reference cells may besignificantly reduced. Reducing the size of the sample cell and itscorresponding cell stem, in turn, reduces the amount of biologicalsubstance used for each experiment.

The following Table 1 represents certain key dimensions of the cellstems and test and reference cells of both the prior art ITC and the ITCof the present invention. As shown in Table 1, the inventive ITC, whichplaces the stirring motor directly on the pipette subassembly, ratherthan positioning it on the calorimeter body, allows a smaller volume oftest substance to be used, thereby reducing the costs associated withconducting calorimeter experiments.

TABLE 1 Prior Art ITC Inventive ITC Stem Interior Diameter 2.6millimeters 2.0 millimeters Cell stem Length 72.0 millimeters 40.5millimeters Cell stem Volume 382.0 microliters 127.0 microliters CellInterior Diameter 22.0 millimeters 11.0 millimeters Cell Thickness 3.7millimeters 2.2 millimeters Cell Volume 1,400 microliters 200microliters

According to one embodiment, the micro titration calorimetry system 200is provided with a pipette guiding mechanism 510 arranged to guide thepipette assembly 220 between and into at least two positions ofoperation. The first position of operation is a pipette washing position580 wherein the titration needle 360 is inserted in a washing apparatus(see FIGS. 5 a-5 c), and the second position of operation is a titrationposition 560 wherein the syringe is inserted into the sample cell 250for calorimetric measurements. In the embodiment of FIG. 2, the guidingmechanism 510 is comprised of a pipette arm 520 that supports thepipette assembly 220, and an essentially vertical guide rod 530. Thepipette arm 520 is moveably attached by a sleeve 540 to the guide rod530, but its motion about the guide rod is restricted by a guide groove550 in the guide rod 530 and a guide pin 560 that protrudes from theinner surface of the sleeve 540 and which fits into the guide groove550. The disclosed guiding mechanism 510 is of rotational type, and thepositions of operation are arranged at equal distance about the centreof rotation of the guide assembly but at different angular positions,wherein movement of the pipette assembly 220 in the vertical directionis restricted to the angular positions of the positions of operation,and wherein rotational movement of the pipette assembly 220 between theangular positions only is permitted when the titration needle 260 isfully retracted from respective positions of operation. FIG. 4 shows theguide mechanism 510 in a position wherein the pipette assembly 220 isfully retracted from the titration position, i.e. from the sample cell250. At this position the guiding mechanism 510 restricts the possiblemovement of the pipette arm 520 to a vertical movement down into thetitration position, or a rotational movement to reach another positionof operation. In a micro titration calorimetry system 200 with twopositions of operation, the washing apparatus 600 may be arranged toallow filling of titrant to the pipette assembly 220 after washing ofthe pipette assembly is completed. Alternatively, filling of the pipettemay be performed through other means, such as a specific filling port inthe syringe or the like.

FIGS. 5 a to 5 c show a schematic perspective view of one embodiment ofa micro titration calorimetry system 200 with a pipette guidingmechanism 510, arranged to guide the pipette assembly 220 to and fromthree different positions of operation 560, 570, 580. According to oneembodiment, the third position of operation is a titrant fillingposition 570, wherein the titration needle 360 is inserted in a titrantsource 590. FIG. 5 a shows the guiding mechanism 510 in a state whereinthe pipette assembly 220 is in the titration position 560. FIG. 5 bshows the guiding mechanism 510 in a state intermediate the titrationposition 560 and the filling position 570. FIG. 5 c shows the guidingmechanism 510 in a state wherein the titration needle 360 of the pipetteassembly 220 is in the filling position 570. As is disclosed in FIGS. 4a to 5 c, the vertical guide grooves 550 associated with a specificposition of operation 560, 570, 580 may be arranged to position thepipette assembly 220 at a predetermined height with respect to theoperation to be performed.

FIG. 6 shows another embodiment of the guide mechanism 510, wherein theguide groove in the guide rod 530 is replaced by a coaxial externalguide sleeve 610 with corresponding guide paths 620 for the guide arm520.

According to one embodiment, the micro titration calorimetry system 200comprises one or more position sensors, not shown, arranged to registerwhen the pipette assembly 220 is positioned at one or more of thepositions of operation 560,570,580, and wherein the associated operationis restricted by the state of the position sensor. A sensor may, e.g. bearranged to register when the pipette assembly 220 is in correctposition for titration 560, and the calorimetry system 200 may bearranged to prevent start of a titration operation unless said sensorconfirms that the pipette assembly 220 is in correct position.

Again referring to the figures, the guiding mechanism 510 ensures properalignment and positioning of pipette assembly 220 in the sample cell 250etc. All elements of the pipette assembly 220 (syringe, needle, paddle,plunger, linear activator, hollow rotor) and the guiding mechanism 510are preferably precisely aligned during the manufacturing process and donot require any additional alignment during the use of the instrument.The pre-set factory alignment significantly improves usability andreliability of the instrument, considerably reduces the amount ofpreparatory time for an experiment, and makes the quality ofmeasurements independent of the user skills. The guiding mechanism 510also enables proper positioning of the pipette assembly 220 in thewashing apparatus 600 for cleaning and drying of the syringe and e.g.for filling the syringe with titrant.

An exemplary method for utilizing the ITC apparatus disclosed hereincomprises the non-manual alignment, both in depth and in breadth, of thesyringe for filling titrant, washing the pipette, and delivering thetitrant to the syringe.

An exemplary method for operating the inventive ITC apparatus comprisesusing the guiding mechanism 510 to position the pipette for filling thesyringe with titrant (see FIG. 5 c). In this position, the end of theneedle with the paddle is placed in the titrant. The plunger of thesyringe is moved from its lower position to its upper position therebyfilling the syringe with the titrant. The sample cell is filled with thesample solution via the cell stem using an auxiliary syringe. Using theguiding mechanism, the pipette is moved to a position for performing anexperiment (see FIG. 5 a). The program for performing the experiment isactivated. Consistent with the program used for the experiment, therotor of the stirring motor rotates the syringe, needle, and paddle withthe assigned speed enabling proper mixing of the reagents. Consistentwith the program used for the experiment (e.g., when a certaintemperature and/or equilibrium are reached), the linear activator movesthe plunger and injects the titrant into the sample solution. Theinjection can be done discretely (step-by-step) or continuously,depending on the program settings. The calorimeter continuously measuresand records the heat release/absorption versus time associated with theinteraction of reagents. The analysis of the results is done accordingto the established algorithm.

Referring again to FIG. 2, the temperature controller 310 is arranged tocontrol the first and second heat pump units 280, 290 in accordance withpredetermined control-modes. The temperature controller 310 isschematically disclosed by functional blocks, and it may either bedesigned as an electronic circuit, as a software in a CPU basedcontroller, or as a combination thereof. In the disclosed embodiment,the thermal controller 310 may be switched between two modes ofoperation:

-   -   Isothermal mode    -   Thermal set mode        by means of a mode select block 630.

In the isothermal mode the thermal controller 310 is arranged to controlthe first heat pump 280 to minimize the temperature difference betweenthe sample cell 250 and the first thermal shield 260. The temperaturedifference between the sample cell 250 and the first thermal shield 260is registered by a temperature sensor arrangement 640 in communicationwith the temperature controller 310. In the disclosed embodiment, thetemperature sensor arrangement 640 is a differential thermocouple thatgives a non-zero signal when there is a temperature difference betweenthe two points of registration, and the output from the thermocouple isconnected to a preamplifier block 650 in the temperature controller 310.The out-put from the preamplifier 650 is directed by the mode selectblock 630 to a first heat pump controller block 660 that controls thefirst heat pump 280 in response to the signal from the preamplifier 650.By this arrangement, the first heat pump controller 660 will strive tocompensate for any thermal difference between the sample cell 250 andthe first thermal shield 260, whereby a compensated adiabatic state isachieved in the calorimeter 210. In order to compensate for minorsystematic drifts in adiabatic behavior, an offset parameter 670 may beapplied on the signal from the preamplifier 650. The offset parameter670 may be set during calibration of the calorimeter 210. In theisothermal mode, the temperature controller 310 is arranged to controlthe second heat pump 290 to keep the second thermal shield 270 at apredetermined temperature, as is defined by a parameter T_(shield). Thecontrol of the second heat pump 290 is performed by a second heat pumpcontroller block 680 that receives an output from a second comparatorblock 690 wherein T_(shield) is compared with the present temperature ofthe second shield registered by a second shield thermal sensor 700. Inthe temperature set mode the temperature controller 310 is arranged tocontrol the first and second heat pump 280, 290 to bring the first andsecond thermal shields 260, 270 to a predetermined temperature, definedby the temperature T_(shield). This mode is mainly used to bring thecalorimeter 210 to the temperature at which the ITC experiments are tobe run. In this mode, a first comparator block 710 compares T_(shield)with the present temperature of the first shield registered by a firstshield thermal sensor 720, and the output from the first comparatorblock 710 is directed by the mode select block 630 to the first heatpump controller 660. The second heat pump 290 is controlled in the samemanner as in the isothermal mode. The thermal set mode may further beused as a stand by mode in order to keep a constant temperature in thecalorimeter 210.

The temperature controller 310 may further comprise a cell temperaturecontroller block 730 for controlling cell heating elements 740 arrangedto heat the sample and the reference cells 240, 250 in accordance with apredetermined temperature set by a parameter T_(cell). The cell heatingelements 740, are mainly used for fine tuning of the temperature of thecells 240, 250.

The parameters T_(shield), T_(cell) and offset may be set via adedicated user interface of the thermal controller, not shown, or e.g.via a calorimeter user interface run on a computer 760 or the like.calorimetric sensors 750 for sensing the temperature difference betweenthe sample cell 250 and reference cell 240 during the ITC experimentsmay be connected to the computer 760, e.g. via a preamplifier 770.

With respect to the above description, it is to be realized that theoptimum dimensional relationships for the parts of the invention, toinclude variations in size, materials, shape, form, function and mannerof operation, assembly and use, are deemed readily apparent and obviousto one skilled in the art, and all equivalent relationships to thoseillustrated in the drawings and described in the specification areintended to be encompassed by the present invention. Therefore, theforegoing is considered as illustrative only of the principles of theinvention. Further, since numerous modifications and changes willreadily occur to those skilled in the art, it is not desired to limitthe invention to the exact construction and operation shown anddescribed, and accordingly, all suitable modifications and equivalentsmay be resorted to, falling within the scope of the invention.

1-21. (canceled)
 22. A micro titration calorimetry system comprising apipette assembly with a titration needle arranged to be inserted into asample cell for supplying titrant, and a pipette guiding mechanismarranged to restrict the movement of the pipette assembly along safepaths to ensure that the titration needle cannot be damaged duringmovement thereof between different positions of operation.
 23. The microtitration calorimetry system of claim 22, wherein the pipette guidingmechanism is arranged to guide the pipette assembly between and into atleast two positions of operation, wherein a first position of operationis a pipette washing position wherein the titration needle is insertedin a washing apparatus, and a second position of operation is atitration position wherein the syringe is inserted into the sample cellfor calorimetric measurements.
 24. The micro titration calorimetrysystem of claim 22, wherein the washing apparatus is arranged to allowfilling of titrant to the pipette assembly after washing of the pipetteassembly is completed.
 25. The micro titration calorimetry system ofclaim 22, further comprising a third position of operation in the formof a titrant filling position, wherein the titration needle is insertedin a titrant source.
 26. The micro titration calorimetry system of claim23, wherein the guiding mechanism is of rotational type, and thepositions of operation are arranged at equal distance about the centreof rotation of the guide assembly but at different angular positions,wherein movement of the pipette assembly in the vertical direction isrestricted to the angular positions of the positions of operation, andwherein rotational movement of the pipette assembly between the angularpositions only is permitted when the needle is fully retracted fromrespective positions of operation.
 27. The micro titration calorimetrysystem of claim 22, further comprising a position sensor arranged toregister when the pipette assembly is positioned at one of the positionsof operation, and wherein the associated operation is restricted by thestate of the position sensor.
 28. A pipette assembly for an isothermaltitration micro calorimetry system, comprising a titration needlearranged to be inserted into a sample cell for supplying titrant, thetitration needle is rotatable with respect to the micro calorimetrysystem and is provided with a stirring paddle arranged to stir fluid inthe sample cell, wherein the pipette assembly comprises a stirring motorfor driving the rotation of the titration needle.
 29. The automaticpipette assembly of claim 28, wherein the stirring motor is a directdrive motor with a hollow rotor arranged concentric with the titrationneedle.
 30. The automatic pipette assembly of claim 28, wherein thestirring motor drives the titration needle for rotation by a rotationtransmission arrangement.
 31. An isothermal titration micro calorimetrysystem with the automatic pipette assembly of claim 28.